RNA-Seq Data Integration and Normalization

To allow for consistent comparison across datasets, all read mapping was carried out using TopHat 1.1.0 [29 (link)] with supplied annotations and the --no-novel-juncs option set, except for the SOLiD datasets, which were only available in a pre-aligned form with mapping by BioScope 1.2.1. All expression estimation and bias correction were done using Cufflinks 0.9.3 with the same annotation and reference sequence as TopHat. In the case of strand-specific libraries, the correct --library-type option was used as per the Cufflinks manual. For the mouse dataset in the NanoString experiment, the RefSeq refGene annotation for assembly NCBI37/mm9 was used, and was downloaded from the UCSC Genome Browser. For all human datasets, the RefSeq refGene annotation for assembly NCBI36/hg18 [30 (link)] was used, and was downloaded from the UCSC Genome Browser. The only filtering was to remove non-chromosomal and 'random' contigs. After quanti cation with Cufflinks, the subset of transcripts with 1-to-1 mappings to the TaqMan qRT-PCR probes were selected (as listed in the supplement to [16 (link)]) to be used in the correlation tests. All yeast datasets used the Ensembl Saccharomyces cerevisiae annotation, release 59, which was downloaded from the Ensembl website [31 (link)]. Mitochondrial, non-coding, and ribosomal RNA sites were masked in the annotation. All remaining transcripts were used in our correlation tests.